Initial experiments have involved rat skeletal muscle acetylcholine receptor (AchR) solubilized by Triton x-100. However, it was found that sonicated (unimellar) phospholipid vesicles are adversely affected by very low Triton concentrations (greater than .005%) below which the receptor also tends to aggregate. Consequently, we have been using sodium deoxycholate solubilized AchR since this detergent can be more easily removed from proteins and begins to affect vesicle permeabilities at higher concentrations (greater than .01%). We have found no evidence of 3H-bungarotoxin-labelled receptor or unlabelled receptor reconstitution into phosphatidyl choline vesicles either in the presence or absence of Ca ion and Mg ion. Using a more purified AchR, that is free of all but trace amounts of detergent, we are currently investigating reconstitution into negatively charged (10% phosphatidyl serine, 90% phosphatidyl choline) sonicated vesicles. However, we have not yet established the appropriate divalent cation levels necessary to promote the initial receptor-vesicle binding interaction. If this approach fails, we will utilize positively charged vesicles. Finally, we are concurrently attempting reconstitution by the slow detergent dialysis method that is used by Dr. Ephriam Racker's laboratory. Following our initial test for successful reconstitution of the AchR, we propose to perform the following experiments: 1) Determination of the morphology of AchR-reconstituted vesicles by freeze-fracture electron microscopy. 2) The investigation of Na 22 ion leakage from proloaded AchR-reconstituted vesicles in the presence of cholinergic agonists and antagonists. 3) Determination of drug binding to reconstituted AchR by equilibrium dialysis. 4) The phospholipid dependence of the above processes.